Field of Invention
This invention relates to a novel, mutated Salmonella enterica serovar Typhimurium that can be used to generate an immune response in porcine and avian, in particular, and in other animals, in general. This immune response helps reduce morbidity and mortality caused by any serovar of Salmonella enterica and reduces transmission, colonization and carriage of any serovar of Salmonella enterica in the animal.
Description of the Prior Art
Bacteria of the genus Salmonella spp. are notorious for their pathogenicity in both man and animal. There are greater than 2,400 Salmonella serovars that are ubiquitous in the environment and can colonize food producing animals and poultry as well as wild animals and birds without causing overt disease. In the U.S. alone, on a yearly basis the number of humans suffering from Salmonella infections exceeds one million cases. In most cases, the infection is caused by contaminated food. Well-known sources of infection are eggs (from both ducks and chickens), products containing eggs, and insufficiently heated poultry, bovine, and porcine meat. The ability to cope with such infections is low, especially in infants, young children, elderly people and immune compromised people. In these groups, the yearly death rate caused by Salmonella infections is high. During the last few decades large-scale animal husbandry has led to an enormous increase in animal density. As a result, an increase is seen in the number of animal infections and, subsequently, in the number of human infections caused by contaminated food. It is clear that animals are the main source of Salmonella infection in humans. This source is very difficult to control for the following reasons. First, Salmonella infections usually cause no serious illness in healthy, full-grown animals; these animals can carry the bacterium for a prolonged period. Carrier animals shed the bacterium in their feces. Thus it is practically impossible for young animals to avoid infection. Second, many Salmonella species colonize several different host species. Some Salmonella species cause primary infections in specific hosts; other Salmonella species are not restrictive at all. As primary infectans, S. enterica serovar Typhi and S. enterica serovar Paratyphi are frequently associated with infection in man whereas S. enterica serovar Dublin is a causative agent of disease in cattle. S. enterica serovar Abortus-equi causes abortion in horses; S. enterica serovar Abortus-ovi causes abortion in sheep. S. enterica serovar Choleraesuis is the cause of lethal diarrhea in young pigs; S. enterica serovar Typhimurium and S. enterica serovar Enteritidis cause salmonellosis in humans, poultry, pigs, cattle and rodents. S. enterica serovar Arizonae causes disease in turkeys, whereas S. enterica serovar Gallinarum causes salmonellosis only in poultry.
Several years ago, greater than 50% of swine production sites (representing 94% of the U.S. swine inventory) tested positive for Salmonella spp. Swine that are Salmonella carriers are a food safety risk for consumers of pork, an animal health risk to non-colonized/uninfected pigs, and an environmental risk caused by fecal shedding of the pathogen into manure that is used as a soil amendment. Also several years ago, approximately 40% of U.S. dairy operations tested positive for Salmonella spp., and approximately 28% of dairy operations tested positive for Salmonella in either bulk tank milk or in-line milk filters by RT-PCR. Further, in a more recent study, approximately 9% of beef cow-calf operations were positive for Salmonella spp.
Clearly there is a need for safe and efficacious vaccines to combat the various Salmonella species not only to prevent disease in animals but also to reduce subclinical infections in food-producing animals that are a source of foodborne disease in humans. Although attenuated Salmonella vaccines are currently available for swine, these vaccines tend to be Salmonella serovar specific and, therefore, do not provide broad protection against many Salmonella serovars. For example, Boehringer Ingelheim Vetmedica sells an avirulent, attenutated S. enterica serovar Choleraesuis vaccine called Enterisol® SC-54 that provides good protection against S. enterica serovar Choleraesuis infections in swine but does not protect against other Salmonella species. Lohmann Animal Health sells AviPro® MEGAN®VAC 1, an attenuated S. enterica serovar Typhimurium that slightly reduces the amount of S. enterica serovar Typhimurium, S. enterica serovar Enteritidis and S. enterica serovar Heidelberg in the organs of young growing chickens and S. enterica serovar Enteritidis colonization of the crop and digestive tract, including the ceca. However, one can improve greatly upon the efficacy of both of these vaccines.
Prior art approaches to generate attenuated Salmonella spp. used a limited set of mutations. See, e.g., Cardenas, et al., Clin. Microbial Rev. 5:328-342 (1992); Chatfield, et al., Vaccine 7:495-498 (1989). These approaches include (i) temperature sensitive mutants (Fahey, et al., Infect. Immun. 1:263-270 (1970)), (ii) mutants defective for aroA, asd, cys, or thy (Galan, et al., Gene 94:29-35 (1990); Hoiseth, et al., Nature 291:238-239 (1981); Robertsson, et al., Infect. Immun. 41:742-750 (1983)), (iii) Δpur and Δdap mutants (Clarke, et al., J. Vet. Res. 51:32-38 (1987); McFarland, et al., Microb. Pathog. 3:129-141 (1987)), (iv) strains defective in carbohydrate metabolism (e.g. ΔgalE (Germanier, et al., Infect. Immun. 4:663-673 (1971); Hone, et al., J. Infect. Dis. 156:167-174 (1987)), (v) lipopolysaccharide synthesis defective strains (e.g., galE, pmi, rfa) (e.g., Nagy, et al., Infect. Immun. 74(10):5914-5925 (2006) and Nagy, et al., Infect. Immun. 72(7):4297-4301 (2004)), and (vi) strains lacking plasmids and mutants lacking virulence genes (e.g., invA). However, the effectiveness of these approaches is doubtful because, at least until now, they have been unable to confer long term immunity against Salmonella infections and have not been effective against a broad range of Salmonella spp.
There remains a need for a mutated Salmonella immunogenic composition capable of inducing high immune responses in animals. It is expected that such a mutated Salmonella immunogenic composition would induce a strong immune response in an animal, be cross-protective against other Salmonella species (or at least other serovars), and reduce colonization and fecal shedding of Salmonella species. Further such a mutated Salmonella immunogenic composition would be attenuated compared to the wild-type Salmonella bacteria. Such an immunogenic composition would limit the colonization of swine and other animals with Salmonella spp. (or at least other serovars), thereby reducing pathogen transmission and the risks to public health, animal health, and the environment. This invention involves a S. enterica serovar Typhimurium containing one or more mutations in rfaH, omrA, omrB, rybB, micA, and/or invR and the use of the mutated S. enterica serovar Typhimurium to generate in animals an immune response cross-reactive to other Salmonella enterica serovars.